Blumea balsamifera Alleviates Rheumatoid Arthritis by Suppressing Synovial Inflammation via the PI3K/AKT Signaling Pathway: From Chemical Profiling to Molecular Mechanism

<b>Background/Objectives</b>: <i>Blumea balsamifera</i> (L.) DC. (Miao Medicine: Diangd vob bvid), a medicinal plant with a long ethnopharmacological history in Southwest China, is widely used by the Miao, Li, and Zhuang ethnic groups to treat rheumatic diseases. While previous studies indicated that the ethyl acetate fraction of <i>B. balsamifera</i> (BBEA) possesses potent anti-inflammatory properties, its specific bioactive material basis and the underlying molecular mechanisms against rheumatoid arthritis (RA) remain elusive. <b>Methods</b>: In this study, an integrated strategy combining chemical profiling, network pharmacology, and experimental validation was employed. First, UPLC-Q-Exactive-MS/MS analysis was conducted to characterize the chemical constituents of BBEA. Subsequently, network pharmacology and molecular docking were utilized to predict potential active compounds and core signaling pathways. Finally, the therapeutic effects and mechanisms were validated in vivo using a collagen-induced arthritis (CIA) rat model and in vitro using lipopolysaccharide (LPS)-induced RAW264.7 macrophages. <b>Results</b>: A total of 25 active constituents, including Genkwanin and Luteolin, were identified in BBEA via UPLC-Q-Exactive-MS/MS. Network pharmacology analysis predicted that the PI3K/AKT signaling pathway is critical for BBEA's anti-RA activity, and molecular docking confirmed strong binding affinities between key components (e.g., Genkwanin) and core targets (SRC, AKT1). In vivo experiments demonstrated that BBEA significantly reduced the Arthritis Index (AI) and paw swelling, reversed weight loss, and ameliorated synovial hyperplasia in CIA rats (<i>p</i> < 0.05). Furthermore, BBEA markedly downregulated the levels of pro-inflammatory cytokines (<i>TNF-α</i>, <i>IL-1β</i>, <i>IL-6</i>, and <i>IL-17</i>) both in serum and synovial tissues. Mechanistically, Western blot analysis verified that BBEA inhibited the phosphorylation of PI3K and AKT in a dose-dependent manner. <b>Conclusions</b>: This study systematically reveals that BBEA alleviates RA symptoms and synovial inflammation primarily by inhibiting the PI3K/AKT signaling pathway. These findings provide a scientific basis for the traditional application of <i>B. balsamifera</i> and suggest BBEA as a promising candidate for RA therapy.